Ly6C+Ly6G− Myeloid‐derived suppressor cells play a critical role in the resolution of acute inflammation and the subsequent tissue repair process after spinal cord injury

Acute inflammation is a prominent feature of central nervous system (CNS) insult and is detrimental to the CNS tissue. Although this reaction spontaneously diminishes within a short period of time, the mechanism underlying this inflammatory resolution remains largely unknown. In this study, we demonstrated that an initial infiltration of Ly6C⁺Ly6G^? immature monocyte fraction exhibited the same characteristics as myeloid‐derived suppressor cells (MDSCs), and played a critical role in the resolution of acute inflammation and in the subsequent tissue repair by using mice spinal cord injury (SCI) model. Complete depletion of Ly6C⁺Ly6G^? fraction prior to injury by anti‐Gr‐1 antibody (clone: RB6‐8C5) treatment significantly exacerbated tissue edema, vessel permeability, and hemorrhage, causing impaired neurological outcomes. Functional recovery was barely impaired when infiltration was allowed for the initial 24 h after injury, suggesting that MDSC infiltration at an early phase is critical to improve the neurological outcome. Moreover, intraspinal transplantation of ex vivo‐generated MDSCs at sites of SCI significantly reduced inflammation and promoted tissue regeneration, resulting in better functional recovery. Our findings reveal the crucial role of an Ly6C⁺Ly6G^? fraction as MDSCs in regulating inflammation and tissue repair after SCI, and also suggests an MDSC‐based strategy that can be applied to acute inflammatory diseases.     

Volatile Flavor Components of Dried Bonito (Katsuobushi) II. From Neutral Fraction

The aqueous extract of dried bonito (Katsuobushi) was distilled under reduced pressure. The resulting distillate with diethyl ether and the extract was separated into acidic, phenolic, basic and neutral fractions. The neutral fraction was further fractionated into ten sub-fractions by silica gel column chromatography. All these sub-fractions were analyzed by gas chromatography and gas chromatography-mass spectrometry.

One hundred and sixty-five compounds were identified and 12 compounds were tentatively identified from the neutral fraction. Among them, 111 compounds were newly identified as flavor components of Katsuobushi.     

Some Physicochemical Properties of Invertase of Candida utilis

This study examined the microbial transformation of carbazole (CZ) by an isolated bacterium that can use CZ as a sole carbon and nitrogen source. The strain identified as Pseudomonas stutzeri produced a large amount of anthranilic acid (AA) from CZ in the medium containing a nonionic surfactant. In dialysis culture using ion-exchange resin, 7.9 g/liter (58mm) of AA was accumulated from 15g/liter (90mm) of CZ and the molar yield of AA reached about 64%.     

Glucosyl Glycerophosphoric Acid and its Polymer Excreted by α-Amylase-forming Bacillus subtilis

α-Amylase formation by washed cell suspensions of Bac. subtilis was found to be accompanied by the excretion of a compound consisting of glucose, glycerol and phosphoric acid. It was excreted as a polymer and a monomer. The former, a kind of teichoic acid, was significantly dominant in quantity when the cells were incubated under the conditions suitable for α-amylase formation. On the other hand, the monomer prevailed when the bacterial cells were under the unfavorable conditions for the enzyme formation.

Both compounds were purified by ion exchange column chromatography. Chemical and enzymatic investigations revealed the following structures: 2-O-α-d-glucopyranosyl-glycerol-3-monophosphoric acid for the monomer, and a polymerized form of the monomer through phosphodiester linkages involving the hydroxyl groups on C3 of the glycerol, for the polymer.     

The Chromatographic Purification of Yeast Invertase by an Ion-Exchange Resin Method and Some Properties of the Enzyme Obtained

The purification of yeast invertase was attempted by application of the chromatographic method using Duolite C-10, a sulfonic acid cation exchange resin. This method was found to be extremely simple in process and significantly effective for the improvement of purity of the enzyme, compared with those other methods reported, hitherto. In the present paper, the procedure of the purification and some properties of the enzyme obtained thereby, are described, and some discussion of the implications is presented.     

Studies on Bacterial Protease

The neutral protease of Bacillus amylosacchariticus was inactivated by low concentrations of several metal-chelating agents and the inactivated enzyme with EDTA restored its activity almost completely by the addition of Zn⁺⁺ or Co⁺⁺ and partially by Fe⁺⁺ or Mn⁺⁺, if these metal ions were added shortly after the EDTA-treatment. The native enzyme was found to contain 0.19% of zinc together with a significant amount of calcium. Parallel increase in specific activity and zinc content of enzyme preparation was observed throughout the purification procedure. The elution pattern of enzyme activity on a CM-cellulose column chromatography also completely coincided with that of protein-bound zinc. A zinc-free inactive enzyme was also reactivated by the addition of zinc or cobalt ions, clearly showing that the neutral protease of B. amylosacchariticus is a zinc mctalloenzyme.     

Some Properties of an Invertase-liberating Enzyme Isolated from Zymolyase

An enzyme which released invertase from cell ghosts of Candida utilis was isolated in an electrophoretically pure state from “Zymolyase.” The molecular weight of the purified enzyme was estimated to be 5.8 × 10⁴, and its isoelectric point was pH 6.9. The enzyme was stable in a pH range from 6.0 to 9.0, and the optimal pH for liberation of invertase from cell ghosts was around 6.0. The activity of the enzyme was competitively inhibited by glucose, mannose, and sucrose. Unlike the starting enzyme preparation, “Zymolyase,” the purified enzyme released invertase without making holes on the surface of the cell ghosts. Various tests were applied, but the specificity of the enzyme was not defined.     

Studies on Urate Oxidase of Candida utilis

Some physical and chemical properties of urate oxidase (EC 1.7.3.3) isolated from the cells of Candida utilis were investigated. The molecular weight was estimated to be 1.2 × 10⁵ by the equilibrium sedimentation and gel filtration methods. The isoelectric point was determined as 5.4 by the method of density electrofocusing. The enzyme showed a slight absorption at 410 mμ, and the absorbancy at this wave length was only 3% of that at 280 mμ. Contrary to urate oxidase from swine liver, the enzyme from yeast contained a negligible amount of copper, but it contained iron of nearly one atom per mole of the enzyme protein. The yeast urate oxidase was not inactivated by some chelators. However, it was easily inactivated with certain heavy metal ions such as Hg²⁺, and the inactivated enzyme was reactivated by the addition of thiols, indicating that the enzyme is a sulfhydryl enzyme. The inactivation of the enzyme with urea, on the other hand, was greatly accelerated by the addition of thiols, and some discussion was added to the results obtained.